Summary of Work: Our previous work has shown that nucleotide excision repair is defective in the Li-Fraumeni syndrome, a heritable cancer prone syndrome associated with increased susceptibility to breast cancer. To explore the possible role of DNA repair mechanisms in sporadically occurring breast cancer, nucleotide excision repair of UV-induced dimers was studied in normal human mammary epithelial tissue and in hormone dependent and independent breast tumor cell lines. Studies of bulk DNA repair reveal a defect in the processing and repair of UV-induced cyclobutane pyrimidine dimers for both estrogen receptor positive and estrogen receptor negative breast cancer lines. Furthermore, results of gene-specific repair experiments performed on both tumor cell lines and normal HMEC, suggest that transcription-coupled repair in the two tumor cell lines is defective. Taken together, these results suggest that defective DNA repair may be important in both heritable and sporadic breast cancer. Increased sensitivity to endogenous DNA damage and/or defective DNA repair of other lesions may be important susceptibility factors in the development of sporadically occurring breast neoplasms as well. Increased levels of oxidative DNA lesions have been observed in human breast tumors, suggesting that oxidative DNA damage may play a crucial role in mammary carcinogenesis. Accumulation of oxidative damage may be a result of increased susceptibility to reactive oxygen species. Alternatively, impaired DNA repair mechanisms may fail to eliminate the oxidative lesions thereby resulting in accumulation of DNA damage and subsequent development of mutations and genetic instability. Due to the high levels of oxidative DNA lesions in breast cancer tissue, it is speculated that DNA repair capacity in breast cancer cells may be diminished. Classically, breast tumorigenesis in women has been associated with the hormonal status and genetic inheritance factors. Traditionally, estrogen has been implicated in breast cancer development. The role of estrogen in DNA damage and repair in human mammary tissue is unknown. New thoughts about its role in breast tumorigenesis center on its possible role on induction of oxidative DNA damage and possibly modulation of DNA repair efficiency. With the recent reports that estrogen may increase DNA damage through its free-radical inducing metabolites, it is even more compelling now to find out whether exposure of human mammary cells to estrogen increases DNA damage or modulates DNA repair mechanisms. Our studies attempt to delineate the role of estrogen on DNA repair in human mammary cells. Thus, the long-term goal of the studies described below is to determine the role of estrogen in DNA repair in human mammary cells. This overall goal will be achieved by testing the following hypotheses: (i) Estrogen modulates DNA repair through transcriptional regulation of DNA repair genes. (ii) Estrogen influences DNA repair at a gene specific level. (iii) Estrogen influences DNA repair through modulation of chromatin architecture. In these studies, H2O2-induced damage will be used to study the effects of estrogen on DNA repair in normal mammary cells and breast cancer cell lines. The efficiency of DNA repair mechanisms as well as hormonal factors may be crucial determinants of mammary carcinogenesis. The results of these experiments may further the understanding of the molecular effects of estrogen on human breast biology in terms of DNA damage and repair.